The physiological functions of the opioid-like receptor ORL1 have been investigated to a great extent since the discovery of its endogenous ligand, nociceptin/Orphanin FQ (N/OFQ). Although it was initially found to be nociceptive, N/OFQ has also been shown to be analgesic under certain experimental conditions. Because N/OFQ is a 17 amino acid peptide, it must be administered directly into the brain or spinal cord, and it degrades relatively quickly. More importantly, there have been no selective antagonists to define the ORL1-mediated actions of N/OFQ. To better understand the actions of N/OFQ and ORL1, a variety of methods will be used to identify compounds that modulate the activities of ORL1. Studies will also be initiated to characterize and modulate the promoter region of proN/OFQ and thus the expression of N/OFQ. These studies indirectly serve the same function, in that the activity of ORL1 will be affected. The avenues to be explored, as reflected in the Specific Aims, include several independent methods to identify non-peptide agonists and antagonists of ORL1. These methods include screening of combinatorial libraries and fungal extracts as well as standard medicinal chemistry. The physiological actions of non-peptide agonists and antagonists will be examined in vitro by their actions on [35S]GTPgammaS binding and cAMP accumulation in CHO cells transfected with ORL1, and in vivo by their effects on antinociception in mice. The promoter region of proN/OFQ will be cloned, sequenced, transfected into an expression vector, and examined to gain a better understanding of the regulation of proN/OFQ transcription. The use of combinatorial libraries or fungal extracts will also be explored to test the hypothesis that small molecules can be identified that might modulate proN/OFQ transcription. Finally, we will test the hypothesis that endogenous substances can be isolated from rat brain that might act as functional ORL1 antagonists. Each of these aims represents a step toward understanding the actions of ORL1 and N/OFQ and exploring into the usefulness of this receptor system for the production of non-addicting analgesics or other therapeutic agents.